Here we outline three projects and summarize the progress. Project 1: The role of protracted cocaine withdrawal in the development of functional and structural plasticity in the NAc We completed a study that investigated the role of protracted cocaine withdrawal by comparing the outcome of short- and long-term cocaine administration in mice. The study involved parallel electrophysiological recordings and morphological studies in the same animals to address the functional and morphological changes induced by cocaine at these synapses. Three main observations were made: 1) The enhancement by cocaine of glutamatergic transmission in the NAc develops with time. A few repeated cocaine injections can trigger the plasticity but the cocaine-induced changes do not express until weeks have elapsed. 2) However, contrary to current views, our data indicate that protracted cocaine withdrawal is not required for the development of plasticity in the NAc, because functional and morphological changes were detected right after a long cocaine treatment that avoided prolonged abstinence. 3) Medium spiny neurons (MSNs) expressing dopamine D1-receptors in the core of the NAc are particularly susceptible to the cocaine-induced plasticity. By revealing a time-dependent plasticity that does not require prolonged abstinence, our findings suggest novel hypotheses about the mechanisms of action of cocaine in the NAc. Project 2: Glutamate evoked calcium signals in spines of medium spiny neurons in the nucleus accumbens and the changes caused by cocaine exposure Dendritic spines in MSNs can receive both glutamatergic and dopaminergic inputs and represent one compartmentalized structure where the integration and modulation among these inputs can occur. This study used electrophysiological recordings in combination with 2-photon laser scanning imaging and uncaging of glutamate to probe glutamate receptor density and composition, as well as local calcium signals at individual spines in MSNs from saline controls and cocaine treated mice. Results show that the AMPAR component of glutamate evoked currents was decreased after cocaine treatment. Interestingly, calcium transients at spines evoked by glutamate receptor activation were larger after the repeated cocaine injections. The increase was completely blocked by NMDAR antagonist and L- and N-type voltage dependent calcium channels blockers indicating NMDAR and L- and N-type voltage dependent calcium channels are responsible for the increased calcium transient. Future experiments will employ pharmacological agents to address the contribution of NR2B containing NMDA receptors to increased glutamate-evoked calcium signals in spines of D1-MSNs shortly after cocaine treatment. Project 3: The development of addictive-like behaviors in mice and associated synaptic changes A major goal of our current research is to understand what is different in the brain of animals that developed addictive behaviors. Addressing this important issue is complicated and progress has been limited by the availability of animal models of addiction. This study develops a mouse model of cocaine self-administration (SA) that identifies addictive-like behaviors by evaluating 3 criteria described in the DSM-IV to define drug dependence: (1) perseverance, shown as difficulty in stopping drug use (2) motivation of cocaine seeking, shown by the effort exerted to obtain the drug, and (3) continued cocaine SA despite adverse consequences. This model is based on a rat SA model described in Deroche-Gamonet et. al. Science (2004). Nave adult mice were trained to nose poke to gain an intravenous infusion of cocaine paired with a cue light. During training, mice learned to discriminate between the active and inactive nose hole and an increase in the rewards/session was observed. When tested, a typical inverted U shaped dose response curve was observed with maximum responses achieved for 1 and 2 mg/kg cocaine doses. Mice were then moved to self-administration (SA) sessions for 40 days, where a wide range of mean daily cocaine intake was found among the mice (5.5 to 23 mg/kg). When screened for the development of addictive-like behaviors, a quarter of the mice (25%) revealed perseverance;19% were highly motivated to obtain cocaine and 19% showed insensitivity to punishment in the SA behavior. Notably, the development of perseverance, high motivation, or insensitivity to adverse consequences was not correlated with the average daily intake. Based on the pattern of drug intake, two distinct groups can be distinguished: steady-takers and binger-takers. Steady-takers acquired reward evenly throughout the session and rewards were spaced apart while binge-takers acquired rewards in clusters, 3 or more and rewards tend to happen at the beginning of the session. These mice have higher mean intake. We have successfully implemented a model of cocaine self administration and we are now investigating the synaptic changes in the NAc associated with the development of addictive like behaviors and different intake patterns.